The binding of copper (Cu2+) and nickel (Ni2+) ions to serum albumin has been studied by 35Cl nuclear magnetic resonance (NMR). The number of high affinity binding sites estimated by the NMR technique agreed well with previous data obtained with an ion-specific electrode. The estimated correlation time of albumin bound Ni2+ suggests that the N-terminal metal ion binding site undergoes more rapid motion than the protein molecule as a whole. Circular dichroism studies showed marked differences in Cu2+ and Ni2+ binding to dog serum albumin as compared to human and bovine serum albumin. Attempts to use 19F NMR to monitor ligand binding to human serum albumin were unsuccessful.